Target: Serine Threonine Protein Phosphatase 1 (PP1) *NCBI Gene # or RefSeq#: 811723 (PP1) *Protein ID (NP or XP #) or Wolbachia#: Pf3D7_1414400 Organism (including strain): Plasmodium falciparum Etiologic Risk Group (see link below): Group B ABSA ranking - 2 Disease Information: Plasmodium falciparum is the predominant causative agent of the disease malaria. Malaria is found predominantly in tropical and subtropical regions but can pose a risk to North America, Europe, and northern Asia [1]. Malaria’s spread is mediated by infected Anopheles mosquitos transmitting P. falciparum during blood meals. From here, the parasite multiplies in the human liver and infects red blood cells. Through blood meals on infected humans by uninfected Anopheles mosquitos, P. falciparum spreads as the cycle of disease continues [1].
-- -or link (or citation) to paper that contains assay information -- links to assay reagents (substrates) pages. --- List cost and quantity of substrate reagents, supplier, and catalog #
Structure (PDB or Homology model) Closest PDB entry if using homology model: 1JK7 -- For Homology Model option: ---- Query Coverage: 98% ---- Max % Identities: 86% ---- % Positives: 94% Molprobity Comparison
Template
Model Current Inhibitors: cyclosporine A, okadaic acid, trifluoperazine Expression Information (has it been expressed in bacterial cells): It has been expressed in E. coli M15 cells. Purification Method: N- terminal affinity tag added to polypeptide. Nickel - chelation chromatography is used for purification of the His tag. Image of protein (PyMol with features delineated and shown separately): Amino Acid Sequence (paste as text only - not as screenshot or as 'code'): MALEIDIDNVISKLIEVRGTRPGKNVNLTENEIKILCLSSREIFLNQPILLELEAPIKIC GDIHGQFYDLLRLFEYGGFPPDANYLFLGDYVDRGKQSLETICLLLAYKIKYPENFFLLR GNHECASINRIYGFYDECKRRYSVKLWKTFIDCFNCLPVAAIIDEKIFCMHGGLSPELNN MEQIRKITRPTDVPDNGLLCDLLWSDPEKEINGWGENDRGVSFTFGQDVVHNFLRKHELD LICRAHQVVEDGYEFFAKRQLVTLFSAPNYCGEFDNAGAMMSVDETLMCSFQILKPVEKK KAAN *length of your protein in Amino Acids: 304 (323 residues in 1JK7) Molecular Weight of your protein in kiloDaltons using the Expasy ProtParam website: 34.9042 kDa Molar Extinction coefficient of your protein at 280 nm wavelength: 33640 (assuming all Cys residues form cystines); 32890 (assuming a;; Cys residues are reduced) TMpred graph Image (http://www.ch.embnet.org/software/TMPRED_form.html). Input your amino acid sequence to it. *CDS Gene Sequence (paste as text only):
TACTTCCAATCCATGGCGCTGGAAATC *GC% Content for gene: 48.1% *CDS Gene Sequence (codon optimized) - copy from output of Primer Design Protocol (paste as text only): TATTTCCAAT CAATGGCACT TGAAATT *GC% Content for gene (codon optimized):
29.6%
*NCBI Gene # or RefSeq#: 811723 (PP1)
*Protein ID (NP or XP #) or Wolbachia#: Pf3D7_1414400
Organism (including strain): Plasmodium falciparum
Etiologic Risk Group (see link below): Group B
ABSA ranking - 2
Disease Information:
Plasmodium falciparum is the predominant causative agent of the disease malaria. Malaria is found predominantly in tropical and subtropical regions but can pose a risk to North America, Europe, and northern Asia [1]. Malaria’s spread is mediated by infected Anopheles mosquitos transmitting P. falciparum during blood meals. From here, the parasite multiplies in the human liver and infects red blood cells. Through blood meals on infected humans by uninfected Anopheles mosquitos, P. falciparum spreads as the cycle of disease continues [1].
P. falciparum infections often progress to fatal forms of cerebral malaria, acute renal failure or acute respiratory distress syndrome [1].
Link to TDR Targets page: http://www.tdrtargets.org/targets/view?gene_id=2475
Link to Gene Database page (NCBI, EuPath databases -e.g. TryTryp, PlasmoDB, etc - or PATRIC, etc.)
http://www.ncbi.nlm.nih.gov/gene/813836
http://plasmodb.org/plasmo/showRecord.do?name=GeneRecordClasses.GeneRecordClass&source_id=PF3D7_1414400&project_id=PlasmoDB
Essentiality of this protein:
Is it a monomer or multimer as biological unit? (make prediction at http://www.ebi.ac.uk/msd-srv/prot_int/pistart.html): Monomeric
Complex of proteins?: 1 amino acid chain with 7 ligands. The structure do not form a complex.
Druggable Target (list number or cite evidence from a paper/database showing druggable in another organism):
The links followed below are tested on P. falciparum
Okadaic acid has articles linking to no inhibition and also a link to one with inhibition: Link to No Inhibition Link to Inhibition IC50: 4 nM
Trifluoperazine Inhibition of PfPP2B
*EC#: 3.1.3.16
Link to BRENDA EC# page: 3.1.3.16
Enzyme Assay information (spectrophotometric, coupled assay ?, reagents): Spectrophotometric pNPP assay
-- link to Sigma (or other company) page for assay (see Sigma links below)
N/A Link is offline as of 10/18 12:40 A.M. :
http://www.sigmaaldrich.com/life-science/metabolomics/enzyme-explorer/learning-center/assay-library/ec-number-iii.html
-- -or link (or citation) to paper that contains assay information
-- links to assay reagents (substrates) pages.
--- List cost and quantity of substrate reagents, supplier, and catalog #
Structure (PDB or Homology model)
Closest PDB entry if using homology model: 1JK7
-- For Homology Model option:
---- Query Coverage: 98%
---- Max % Identities: 86%
---- % Positives: 94%
Molprobity Comparison
Template
Model
Current Inhibitors: cyclosporine A, okadaic acid, trifluoperazine
Expression Information (has it been expressed in bacterial cells): It has been expressed in E. coli M15 cells.
Purification Method: N- terminal affinity tag added to polypeptide. Nickel - chelation chromatography is used for purification of the His tag.
Image of protein (PyMol with features delineated and shown separately):
Amino Acid Sequence (paste as text only - not as screenshot or as 'code'):
MALEIDIDNVISKLIEVRGTRPGKNVNLTENEIKILCLSSREIFLNQPILLELEAPIKIC
GDIHGQFYDLLRLFEYGGFPPDANYLFLGDYVDRGKQSLETICLLLAYKIKYPENFFLLR
GNHECASINRIYGFYDECKRRYSVKLWKTFIDCFNCLPVAAIIDEKIFCMHGGLSPELNN
MEQIRKITRPTDVPDNGLLCDLLWSDPEKEINGWGENDRGVSFTFGQDVVHNFLRKHELD
LICRAHQVVEDGYEFFAKRQLVTLFSAPNYCGEFDNAGAMMSVDETLMCSFQILKPVEKK
KAAN
*length of your protein in Amino Acids: 304 (323 residues in 1JK7)
Molecular Weight of your protein in kiloDaltons using the Expasy ProtParam website: 34.9042 kDa
Molar Extinction coefficient of your protein at 280 nm wavelength: 33640 (assuming all Cys residues form cystines); 32890 (assuming a;; Cys residues are reduced)
TMpred graph Image (http://www.ch.embnet.org/software/TMPRED_form.html). Input your amino acid sequence to it.
*CDS Gene Sequence (paste as text only):
TACTTCCAATCCATGGCGCTGGAAATC
*GC% Content for gene: 48.1%
*CDS Gene Sequence (codon optimized) - copy from output of Primer Design Protocol (paste as text only):
TATTTCCAAT CAATGGCACT TGAAATT
*GC% Content for gene (codon optimized):
29.6%
Primer design results for pNIC-Bsa4 cloning (list seqeunces of all of your ~40 nt long primers):
(link to DNA Works output text file)
Primer design results for 'tail' primers (this is just 2 sequences):
Forward
5' TACTTCCAATCCATGGCGCTGGAAATC 3'
Reverse
5' TATCCACCTTTACTGTTAGTTCGC 3'